Cepacidine A, a novel antifungal antibiotic produced by Pseudomonas cepacia. II. Physico-chemical properties and structure elucidation.

Abstract

Cepacidine A is a novel glycopeptide with a potent antifungal activity, which is produced by Pseudomonas cepacia AF 2001. Its molecular weight was determined by FAB-MS(m/z 1215). The compound is comprised of glycine (1), serine (2), 2,4-diaminobutyric acid (1), aspartic acid (1), jS-hydroxy tyrosine (1), /Miydroxy asparagine (1), xylose (1) and 5,7-dihydroxy-3,9-diaminooctadecanoic acid (1). Unfortunately, cepacidine A is a mixture of A± and A2, either of which is barely distinguishable. Cepacidine A2 includes asparagine (1) instead of /?-hydroxy asparagine (1) of cepacidine Av The MSdata and the NOESY, TOCSYand HMBCspectra show that cepacidine A is a cyclic peptide and xylose is connected to 5,7-dihydroxy-3,9-diaminooctadecanoic acid.Cepacidine A is a new metabolite of Pseudomonas cepacia AF 2001 discovered in the author's screening program for antifungal substances. Its discovery, taxonomy, fermentation, isolation and biological properties were reported in the previous paper.1} This paper describes the physico-chemical properties and structure elucidation of cepacidine A. (Fig. 1) Physico-chemical Properties The physico-chemical properties of cepacidine A is summarized in Table 1. Cepacidine A is isolated as white powders. The melting point is ranged between 210°C and 214°C. Cepacidine A is insoluble in ethyl acetate, hexane, ether and benzene, and is hardly soluble in water, methanol, ethanol, isopropanol, butanol and acetone, while soluble in DMSO, alkali aqueous solution and acidic aqueous solution. 50% aqueous solution of alcohol increases solubility. Cepacidine A shows positive color reactions to aniline and ninhydrin reagents. The Rf value of cepacidine A on silica gel, TLC, developed with n-butanol -acetic acid-water (3 : 1 : 1) was 0.18. However, the Rfvalues ofcepacidine A on silica gel, TLC, developed with isopropanol-water-saturated aqueous ammonia (4 : 1 : 2), were separated as 0.53 and 0.58 so that the compound having the Rf value of 0.53 was named cepacidine Al5 and the compound having the Rf value of 0.58 was named cepacidine A2. Since cepacidine Ax and cepacidine A2 each were barely obtainable by using prep-HPLC, it was unfortunate, that the mixture, cepacidine A, was used for all spectrometric analysis with the exception of TLCand amino acid which were analyzed by HPLC.The mixture has the 9: 1 ratio of cepacidine At and cepacidine A2. The UVspectrum of cepacidine A dissolved in water showed two maximumabsorption peaks at 232 nm and 274 nm, and the spectrum in DMSO showed only one peak at 278nm. Cepacidine A is very stable in an aqueous solution ranging between pH 2 and pH 1 1, and it is unstable in aqueous solution above pH ll.5, and it loses its antifungal activities readily. The molecular formula of cepacidine Ax was determined to be C^HgsO^Ni! by HRFAB-MS, 13C NMR and elemental analysis (calcd: C 51.4, H 7.0, N 12.7, O 29.0; found: C 51.5, H 8.0, N ll.0, O 29.5),

A comprehensive review of glycosylated bacterial natural products

“…While most members in this group display antifungal activity (hassillidins from Hassallia , 1093,1094 occidiofungins 1095 and burkholdine 1096 from Burkholderia , cepacidines from Pseudomonas , 1097,1098 herbicolins from Erwinia 1099,1100 and W-10β from Aeromonas 1101 ), the ramoplanins from Actinoplanes stand out as agents recently approved by the FDA to treat C. difficile (Fig. 49).…”

A Burkholderia cepacia complex non-ribosomal peptide-synthesized toxin is hemolytic and required for full virulence

“…These bacteria embody a diverse array of metabolic capabilities, among them environmental contaminant degradation, 1 , 2 fungal phytopathogen suppression 3 - 7 and nitrogen fixation 8 . For these reasons, the Bcc has been identified as a powerful class of bioremediation and plant growth promoting agents 9 .…”

“…Known secondary metabolite toxins produced by the Bcc include the broad-spectrum antifungal agent “pyrrolnitrin,” 29 a group of antifungal and antitumor agents deemed “cepafungins,” 30 hemolytic peptides named “cepalysins,” 3 related antifungal compounds “cepacidine A1 and A2” 7 , 31 and the recently described antifungal cyclic peptide “occidiofungin” or “burkholdine,” 32 - 34 which may be similar to molecules previously isolated from Bcc strains as “xylocandins.” 35 , 36 While these latter compounds show a broad range of activity against fungi, in this manuscript, we identify through random and directed mutagenesis a gene cluster in Burkholderia vietnamiensis strain DBO1 37 that is homologous to the gene cluster that synthesizes occidofungin/burkholdine, and demonstrate that this gene product is also strikingly β-hemolytic. This study also examines the prevalence of this gene cluster within the Bcc and the lethality of this compound in an invertebrate infection model.…”

Antimicrobial Cyclic Peptides for Plant Disease Control

“…3) are glycopeptides produced by Pseudomonas cepacia (Lee et al, 1994; Lim et al, 1994). The MICs for cepacidine A range from 0.098 to 0.391 mg/ml against Aspergillus niger , F. oxysporum , and Rhizopus stolonifer .…”